A major aim of the Integrative Immunobiology Unit is to decipher gene expression programs that direct cell fates in the hematopoietic and immune systems, since perturbations to their genetic program underlie many diseases such as cancer, immunodeficiency, autoimmunity, allergy and infectious diseases. We seek knowledge that will provide insights for understanding these diseases and the parameters for effective immune responses. For a tractable model system to prototype our approaches, we are taking advantage of CD4+ T helper cell differentiation since many research tools are available for this system and much is already known about its core transcriptional program and signaling pathways. However, relatively little is known about post-transcriptional control of gene expression and the roles of non-coding RNAs in immunity. One major focus of this project is a class of endogenous small untranslated RNAs, called microRNAs (miRNAs), that partner with Argonaute (Ago) proteins to form effector RNA-induced silencing complexes (RISCs) that recognize cognate mRNA targets and reduce their expression post-transcriptionally. It became evident that miRNAs are required for B and T lymphocyte differentiation when miRNA biogenesis was blocked by conditional ablation of the miRNA processing enzyme Dicer in genetically engineered mouse models. Recently, we have expanded our research to identify novel long non-coding RNAs (lncRNAs) that are expressed in lymphocytes using a technique called RNA-seq and will endeavor to determine their roles in immunity. Previously, we successfully combined mouse genetics and genomics to systematically determine the impact of miRNAs on the transcriptome. We will continue with this approach to integrate miRNAs and lncRNAs into maps of regulatory networks that orchestrate gene expression in lymphocytes. To accomplish our goals we are establishing state-of-the-art genomic methods enabled by massively parallel sequencing. A typical deep sequencing experiment generates in the order of 100 million reads, that can only be analyzed using sophisticated computational methods. Although, some tools exist for analyses we had to develop some custom in-house source codes for certain applications. We routinely perform deep sequencing for a variety of genomic applications (eg. RNA-seq, ChIP-seq) with the goal of understanding how cells in the immune system dynamically interpret the genetic code within their DNA in order to fulfill their biological destinies. This past year, we have been busy generating, analyzing and mining our data; we formulated a number of interesting hypotheses; and we are intensively testing them experimentally in the laboratory. We surmounted a number of technical challenges and are currently concluding some of our work for publication. In 2012, we reported that the let-7 family of microRNAs are expressed by adult bone marrow hematopoietic progenitors but not their fetal counterparts (Science 335:1195-1200).